Dr. Banik, my collaborator, has isolated and purified calcium activated neutral proteinase (CANP II) from bovine myelin. Human myelin basic protein, component 1 (HBP-1) was digested by CANP, and the peptides were separated on HPLC into pooled fractions. Since the sequence of HBP is known, N- and C-terminal analyses yielded the sequence of the peptides and, therefore, the eleven cleavage sites of CANP. The major site was Val-(94)-Thr(95). Presently, CANP peptides of guinea pig, component 1, which could be encephalitogenic are being purified by HPLC. Then, the purified CANP peptides will be tested for their encephalitogenic activity in Lewis rats. Multicatalytic proteinase complex (MPC) from bovine brain has also been isolated and purified by Dr. Banik. MPC also digests HBP-1 but at a much lower rate. Human myelin basic protein (HBP) occurs in multiple forms. Three of these isoforms have been highly purified - HBP-1 (unmodified BP), 17.2kDa HBP (missing residues 106-116) and HBP-3pT98 (phosphorylated on Thr(98)). When compared with circular dichroism (CD) studies done on HBP-1, the CD studies of 17.2kDa HBP showed about 9% increase in ordered structure and those of HBP3pT98 about a 12% increase. Both modifications of HBP-1, the deletion of residues 106-116 and the addition of a single phosphate promoted secondary structure, probably an increase of ~-structure. The proliferative response of T-cell lymphocytes (TCL) specific for HBP (87-106) to HBP-1, 17.2kDa HBP and HBP 3pT98 was studied by Dr. Voskuhl, my collaborator. All three HBP's were recognized by the TCLs.